Bottom Line:
Twenty isolates were subjected to taxonomic characterization using both ascospore SEM micromorphology and two independent DNA loci sequencing.All the isolates produced neoechinulin A, 17 isolates were positive for cladosporin and echinulin, and 18 were positive for neoechinulin B.Physcion and preechinulin were detected in a minor proportion of the isolates.

ABSTRACTXerophilic fungal species of the genus Aspergillus are economically highly relevant due to their ability to grow on low water activity substrates causing spoilage of stored goods and animal feeds. These fungi can synthesize a variety of secondary metabolites, many of which show animal toxicity, creating a health risk for food production animals and to humans as final consumers, respectively. Animal feeds used for rabbit, chinchilla and rainbow trout production in Argentina were analysed for the presence of xerophilic Aspergillus section Aspergillus species. High isolation frequencies (>60%) were detected in all the studied rabbit and chinchilla feeds, while the rainbow trout feeds showed lower fungal charge (25%). These section Aspergillus contaminations comprised predominantly five taxa. Twenty isolates were subjected to taxonomic characterization using both ascospore SEM micromorphology and two independent DNA loci sequencing. The secondary metabolite profiles of the isolates were determined qualitatively by HPLC-MS. All the isolates produced neoechinulin A, 17 isolates were positive for cladosporin and echinulin, and 18 were positive for neoechinulin B. Physcion and preechinulin were detected in a minor proportion of the isolates. This is the first report describing the detailed species composition and the secondary metabolite profiles of Aspergillus section Aspergillus contaminating animal feeds.

toxins-07-03512-f002: Phylogenetic trees generated by Maximum Likelihood analysis showing the relationship of 18 type strains of Aspergillus section Aspergillus species and the 20 isolates obtained from animal feeds and primary raw materials. Three based on (a) the ITS region; (b) the beta-tubulin gene fragment; and (c) the concatenated analysis of both. Penicillium steckii (NRRL 35625) was used as outgroup. The thickened lines represent lineages with >90% bootstrap values. The Bootstrap analyses were performed with 1000 replications. T Ex-type strain TT Ex-teleotype strain.

Mentions:
The maximum likelihood analysis based on the ITS sequences (Figure 2a) resulted in very weak resolution of the Aspergillus section Aspergillus at species level. All the 20 studied isolates located within two major multi-species terminal clades with strong bootstrap support but none of them could be identified at species level. One of the clades included both the 11 studied isolates and all the species of A. glaucus and A. ruber clades, and A. cibarius. Within this multispecies clade, four of the studied isolates formed part of a moderately supported internal subclade. However, this subclade was neither conspecific nor included both A. ruber and A. appendiculatus. The second major multispecies clade consisted of the rest of the nine studied isolates and the species known to belong to the A. chevalieri clade of the section.

toxins-07-03512-f002: Phylogenetic trees generated by Maximum Likelihood analysis showing the relationship of 18 type strains of Aspergillus section Aspergillus species and the 20 isolates obtained from animal feeds and primary raw materials. Three based on (a) the ITS region; (b) the beta-tubulin gene fragment; and (c) the concatenated analysis of both. Penicillium steckii (NRRL 35625) was used as outgroup. The thickened lines represent lineages with >90% bootstrap values. The Bootstrap analyses were performed with 1000 replications. T Ex-type strain TT Ex-teleotype strain.

Mentions:
The maximum likelihood analysis based on the ITS sequences (Figure 2a) resulted in very weak resolution of the Aspergillus section Aspergillus at species level. All the 20 studied isolates located within two major multi-species terminal clades with strong bootstrap support but none of them could be identified at species level. One of the clades included both the 11 studied isolates and all the species of A. glaucus and A. ruber clades, and A. cibarius. Within this multispecies clade, four of the studied isolates formed part of a moderately supported internal subclade. However, this subclade was neither conspecific nor included both A. ruber and A. appendiculatus. The second major multispecies clade consisted of the rest of the nine studied isolates and the species known to belong to the A. chevalieri clade of the section.

Bottom Line:
Twenty isolates were subjected to taxonomic characterization using both ascospore SEM micromorphology and two independent DNA loci sequencing.All the isolates produced neoechinulin A, 17 isolates were positive for cladosporin and echinulin, and 18 were positive for neoechinulin B.Physcion and preechinulin were detected in a minor proportion of the isolates.

ABSTRACTXerophilic fungal species of the genus Aspergillus are economically highly relevant due to their ability to grow on low water activity substrates causing spoilage of stored goods and animal feeds. These fungi can synthesize a variety of secondary metabolites, many of which show animal toxicity, creating a health risk for food production animals and to humans as final consumers, respectively. Animal feeds used for rabbit, chinchilla and rainbow trout production in Argentina were analysed for the presence of xerophilic Aspergillus section Aspergillus species. High isolation frequencies (>60%) were detected in all the studied rabbit and chinchilla feeds, while the rainbow trout feeds showed lower fungal charge (25%). These section Aspergillus contaminations comprised predominantly five taxa. Twenty isolates were subjected to taxonomic characterization using both ascospore SEM micromorphology and two independent DNA loci sequencing. The secondary metabolite profiles of the isolates were determined qualitatively by HPLC-MS. All the isolates produced neoechinulin A, 17 isolates were positive for cladosporin and echinulin, and 18 were positive for neoechinulin B. Physcion and preechinulin were detected in a minor proportion of the isolates. This is the first report describing the detailed species composition and the secondary metabolite profiles of Aspergillus section Aspergillus contaminating animal feeds.